title: Complete disassociation of adult pancreas into viable single cells through cold trypsin-EDTA digestion.
abstract: The in vitro isolation and analysis of pancreatic stem/progenitor cells are necessary for understanding their properties and function; however, the preparation of high-quality single-cell suspensions from adult pancreas is prerequisite. In this study, we applied a cold trypsin-ethylenediaminetetraacetic acid (EDTA) digestion method to disassociate adult mouse pancreata into single cells. The yield of single cells and the viability of the harvested cells were much higher than those obtained via the two commonly used warm digestion methods. Flow cytometric analysis showed that the ratio of ductal or BCRP1-positive cells in cell suspensions prepared through cold digestion was consistent with that found in vivo. Cell culture tests showed that pancreatic epithelial cells prepared by cold digestion maintained proliferative capacity comparable to those derived from warm collagenase digestion. These results indicate that cold trypsin-EDTA digestion can effectively disassociate an adult mouse pancreas into viable single cells with minimal cell loss, and can be used for the isolation and analysis of pancreatic stem/progenitor cells.
authors.affiliation: Laboratory of Animal Development Biology, College of Life Science, Northeast Forestry University, Harbin 150040, China.
authors.forename: Dan
authors.initials: D
authors.lastname: Li
authors.forename: Shi-Yun
authors.initials: SY
authors.lastname: Peng
authors.forename: Zhen-Wu
authors.initials: ZW
authors.lastname: Zhang
authors.forename: Rui-Cheng
authors.initials: RC
authors.lastname: Feng
authors.forename: Lu
authors.initials: L
authors.lastname: Li
authors.forename: Jie
authors.initials: J
authors.lastname: Liang
authors.forename: Sheng
authors.initials: S
authors.lastname: Tai
authors.forename: Chun-Bo
authors.initials: CB
authors.lastname: Teng
mesh_terms: D000818:Animals; D002452:Cell Count; D018929:Cell Culture Techniques; D049109:Cell Proliferation; D002469:Cell Separation; D002470:Cell Survival; D017364:Collagenases / Q000737:chemistry; D004492:Edetic Acid / Q000737:chemistry*; D005434:Flow Cytometry; D051379:Mice; D008856:Microscopy, Fluorescence; D010179:Pancreas / Q000201:enzymology / Q000378:metabolism / Q000473:pathology*; D013234:Stem Cells / Q000166:cytology; D014357:Trypsin / Q000737:chemistry*
chemical_list: D004492:Edetic Acid; D014357:Trypsin; D017364:Collagenases